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Hallmarks of the developmental cycle of the obligate intracellular pathogenic bacterium Chlamydia are the primary differentiation of the infectious elementary body (EB) into the proliferative reticulate body (RB) and the secondary differentiation of RBs back into EBs. The mechanisms regulating these transitions remain unclear. In this report, we developed an effective novel strategy termed dependence on plasmid-mediated expression (DOPE) that allows for the knockdown of essential genes in Chlamydia. We demonstrate that GrgA, a Chlamydia-specific transcription factor, is essential for the secondary differentiation and optimal growth of RBs. We also show that GrgA, a chromosome-encoded regulatory protein, controls the maintenance of the chlamydial virulence plasmid. Transcriptomic analysis further indicates that GrgA functions as a critical regulator of all three sigma factors that recognize different promoter sets at developmental stages. The DOPE strategy outlined here should provide a valuable tool for future studies examining chlamydial growth, development, and pathogenicity.

Kaposi’s sarcoma herpesvirus (KSHV) ORF34 plays a significant role as a component of the viral pre-initiation complex (vPIC), which is indispensable for late gene expression across beta- and gammaherpesviruses. Although the key role of ORF34 within the vPIC and its function as a hub protein have been recognized, further clarification regarding its specific contribution to vPIC functionality and interactions with other components is required. This study employed a deep learning algorithm-assisted structural model of ORF34, revealing highly conserved amino acid residues across human beta- and gammaherpesviruses localized in structured domains. Thus, we engineered ORF34 alanine-scanning mutants by substituting conserved residues with alanine. These mutants were evaluated for their ability to interact with other vPIC factors and restore viral production in cells harboring the ORF34-deficient KSHV-BAC. Our experimental results highlight the crucial role of the four cysteine residues conserved in ORF34: a tetrahedral arrangement consisting of a pair of C-Xn-C consensus motifs. This suggests the potential incorporation of metal cations in interacting with ORF24 and ORF66 vPIC components, facilitating late gene transcription, and promoting overall virus production by capturing metal cations. In summary, our findings underline the essential role of conserved cysteines in KSHV ORF34 for effective vPIC assembly and viral replication, thereby enhancing our understanding of the complex interplay between the vPIC components. IMPORTANCE The initiation of late gene transcription is universally conserved across the beta- and gammaherpesvirus families. This process employs a viral pre-initiation complex (vPIC), which is analogous to a cellular PIC. Although KSHV ORF34 is a critical factor for viral replication and is a component of the vPIC, the specifics of vPIC formation and the essential domains crucial for its function remain unclear. Structural predictions suggest that the four conserved cysteines (C170, C175, C256, and C259) form a tetrahedron that coordinates the metal cation. We investigated the role of these conserved amino acids in interactions with other vPIC components, late gene expression, and virus production to demonstrate for the first time that these cysteines are pivotal for such functions. This discovery not only deepens our comprehensive understanding of ORF34 and vPIC dynamics but also lays the groundwork for more detailed studies on herpesvirus replication mechanisms in future research. [ABSTRACT FROM AUTHOR]

Background: Organisms frequently experience environmental stresses that occur in predictable patterns and combinations. For wild Saccharomyces cerevisiae yeast growing in natural environments, cells may experience high osmotic stress when they first enter broken fruit, followed by high ethanol levels during fermentation, and then finally high levels of oxidative stress resulting from respiration of ethanol. Yeast have adapted to these patterns by evolving sophisticated "cross protection" mechanisms, where mild 'primary' doses of one stress can enhance tolerance to severe doses of a different 'secondary' stress. For example, in many yeast strains, mild osmotic or mild ethanol stresses cross protect against severe oxidative stress, which likely reflects an anticipatory response important for high fitness in nature. Results: During the course of genetic mapping studies aimed at understanding the mechanisms underlying natural variation in ethanol-induced cross protection against H2O2, we found that a key H2O2 scavenging enzyme, cytosolic catalase T (Ctt1p), was absolutely essential for cross protection in a wild oak strain. This suggested the absence of other compensatory mechanisms for acquiring H2O2 resistance in that strain background under those conditions. In this study, we found surprising heterogeneity across diverse yeast strains in whether CTT1 function was fully necessary for acquired H2O2 resistance. Some strains exhibited partial dispensability of CTT1 when ethanol and/or salt were used as mild stressors, suggesting that compensatory peroxidases may play a role in acquired stress resistance in certain genetic backgrounds. We leveraged global transcriptional responses to ethanol and salt stresses in strains with different levels of CTT1 dispensability, allowing us to identify possible regulators of these alternative peroxidases and acquired stress resistance in general. Conclusions: Ultimately, this study highlights how superficially similar traits can have different underlying molecular foundations and provides a framework for understanding the diversity and regulation of stress defense mechanisms. [ABSTRACT FROM AUTHOR]

Abstract Background Organisms frequently experience environmental stresses that occur in predictable patterns and combinations. For wild Saccharomyces cerevisiae yeast growing in natural environments, cells may experience high osmotic stress when they first enter broken fruit, followed by high ethanol levels during fermentation, and then finally high levels of oxidative stress resulting from respiration of ethanol. Yeast have adapted to these patterns by evolving sophisticated “cross protection” mechanisms, where mild ‘primary’ doses of one stress can enhance tolerance to severe doses of a different ‘secondary’ stress. For example, in many yeast strains, mild osmotic or mild ethanol stresses cross protect against severe oxidative stress, which likely reflects an anticipatory response important for high fitness in nature. Results During the course of genetic mapping studies aimed at understanding the mechanisms underlying natural variation in ethanol-induced cross protection against H2O2, we found that a key H2O2 scavenging enzyme, cytosolic catalase T (Ctt1p), was absolutely essential for cross protection in a wild oak strain. This suggested the absence of other compensatory mechanisms for acquiring H2O2 resistance in that strain background under those conditions. In this study, we found surprising heterogeneity across diverse yeast strains in whether CTT1 function was fully necessary for acquired H2O2 resistance. Some strains exhibited partial dispensability of CTT1 when ethanol and/or salt were used as mild stressors, suggesting that compensatory peroxidases may play a role in acquired stress resistance in certain genetic backgrounds. We leveraged global transcriptional responses to ethanol and salt stresses in strains with different levels of CTT1 dispensability, allowing us to identify possible regulators of these alternative peroxidases and acquired stress resistance in general. Conclusions Ultimately, this study highlights how superficially similar traits can have different underlying molecular foundations and provides a framework for understanding the diversity and regulation of stress defense mechanisms.

Abstract Background Goose, descendants of migratory ancestors, have undergone extensive selective breeding, resulting in their remarkable ability to accumulate fat in the liver and exhibit a high tolerance for significant energy intake. As a result, goose offers an excellent model for studying obesity, metabolic disorders, and liver diseases in mammals. Although the impact of the three-dimensional arrangement of chromatin within the cell nucleus on gene expression and transcriptional regulation is widely acknowledged, the precise functions of chromatin architecture reorganization during fat deposition in goose liver tissues still need to be fully comprehended. Results In this study, geese exhibited more pronounced changes in the liver index and triglyceride (TG) content following the consumption of the high-fat diet (HFD) than mice without significant signs of inflammation. Additionally, we performed comprehensive analyses on 10 goose liver tissues (5 HFD, 5 normal), including generating high-resolution maps of chromatin architecture, conducting whole-genome gene expression profiling, and identifying H3K27ac peaks in the livers of geese and mice subjected to the HFD. Our results unveiled a multiscale restructuring of chromatin architecture, encompassing Compartment A/B, topologically associated domains, and interactions between promoters and enhancers. The dynamism of the three-dimensional genome architecture, prompted by the HFD, assumed a pivotal role in the transcriptional regulation of crucial genes. Furthermore, we identified genes that regulate chromatin conformation changes, contributing to the metabolic adaptation process of lipid deposition and hepatic fat changes in geese in response to excessive energy intake. Moreover, we conducted a cross-species analysis comparing geese and mice exposed to the HFD, revealing unique characteristics specific to the goose liver compared to a mouse. These chromatin conformation changes help elucidate the observed characteristics of fat deposition and hepatic fat regulation in geese under conditions of excessive energy intake. Conclusions We examined the dynamic modifications in three-dimensional chromatin architecture and gene expression induced by an HFD in goose liver tissues. We conducted a cross-species analysis comparing that of mice. Our results contribute significant insights into the chromatin architecture of goose liver tissues, offering a novel perspective for investigating mammal liver diseases.

Background: Goose, descendants of migratory ancestors, have undergone extensive selective breeding, resulting in their remarkable ability to accumulate fat in the liver and exhibit a high tolerance for significant energy intake. As a result, goose offers an excellent model for studying obesity, metabolic disorders, and liver diseases in mammals. Although the impact of the three-dimensional arrangement of chromatin within the cell nucleus on gene expression and transcriptional regulation is widely acknowledged, the precise functions of chromatin architecture reorganization during fat deposition in goose liver tissues still need to be fully comprehended. Results: In this study, geese exhibited more pronounced changes in the liver index and triglyceride (TG) content following the consumption of the high-fat diet (HFD) than mice without significant signs of inflammation. Additionally, we performed comprehensive analyses on 10 goose liver tissues (5 HFD, 5 normal), including generating high-resolution maps of chromatin architecture, conducting whole-genome gene expression profiling, and identifying H3K27ac peaks in the livers of geese and mice subjected to the HFD. Our results unveiled a multiscale restructuring of chromatin architecture, encompassing Compartment A/B, topologically associated domains, and interactions between promoters and enhancers. The dynamism of the three-dimensional genome architecture, prompted by the HFD, assumed a pivotal role in the transcriptional regulation of crucial genes. Furthermore, we identified genes that regulate chromatin conformation changes, contributing to the metabolic adaptation process of lipid deposition and hepatic fat changes in geese in response to excessive energy intake. Moreover, we conducted a cross-species analysis comparing geese and mice exposed to the HFD, revealing unique characteristics specific to the goose liver compared to a mouse. These chromatin conformation changes help elucidate the observed characteristics of fat deposition and hepatic fat regulation in geese under conditions of excessive energy intake. Conclusions: We examined the dynamic modifications in three-dimensional chromatin architecture and gene expression induced by an HFD in goose liver tissues. We conducted a cross-species analysis comparing that of mice. Our results contribute significant insights into the chromatin architecture of goose liver tissues, offering a novel perspective for investigating mammal liver diseases. [ABSTRACT FROM AUTHOR]

Extracellular proteases, such as chitinases secreted by Arthrobotrys oligospora (A. oligospora), play a crucial role in the process of nematode infection. However, post-transcriptional regulation of gene expression involving microRNAs (miRNAs) in A. oligospora remains scarcely described. Hereto, transcriptome sequencing was carried out to analyze the expression profiles of chitin-responsive miRNAs in A. oligospora. Based on the RNA-seq data, the differential expression of miRNAs (DEmiRNAs) in response to chitin was screened, identified and characterized in A. oligospora. Meanwhile, the potential target genes were predicted by the online tools miRanda and Targetscan, respectively. Furthermore, the interaction of DEmiRNA with it’s target gene was validated by a dual-luciferase reporter assay system. Among 85 novel miRNAs identified, 25 miRNAs displayed significant differences in expression in A. oligospora in response to chitin. Gene Ontology (GO) analysis showed that the potential genes targeted by DEmiRNAs were enriched in the biological processes such as bio-degradation, extracellular components and cell cycle. KEGG analysis revealed that the target genes were mainly involved in Hippo, carbon and riboflavin metabolic pathway. Outstandingly, chitinase AOL_s00004g379, which is involved in the hydrolysis metabolic pathway of chitin, was confirmed to be a target gene of differential miR_70. These findings suggest that chitin-responsive miRNAs are involved in the regulation of cell proliferation, predator hyphae growth and chitinase expression through the mechanisms of post-transcriptional regulation, which provides a new perspective to the molecular mechanisms underlying miRNAs-mediated control of gene expression in A. oligospora. [ABSTRACT FROM AUTHOR]

The functional performance of immune cells relies on a complex transcriptional regulatory network. The three-dimensional structure of chromatin can affect chromatin status and gene expression patterns, and plays an important regulatory role in gene transcription. Currently available techniques for studying chromatin spatial structure include chromatin conformation capture techniques and their derivatives, chromatin accessibility sequencing techniques, and others. Additionally, the recently emerged deep learning technology can be utilized as a tool to enhance the analysis of data. In this review, we elucidate the definition and significance of the three-dimensional chromatin structure, summarize the technologies available for studying it, and describe the research progress on the chromatin spatial structure of dendritic cells, macrophages, T cells, B cells, and neutrophils. [ABSTRACT FROM AUTHOR]

Theiler's murine encephalomyelitis virus (TMEV) infection has been used as a mouse model for two virus-induced organ-specific immune-mediated diseases. TMEV-induced demyelinating disease (TMEV-IDD) in the central nervous system (CNS) is a chronic inflammatory disease with viral persistence and an animal model of multiple sclerosis (MS) in humans. TMEV infection can also cause acute myocarditis with viral replication and immune cell infiltration in the heart, leading to cardiac fibrosis. Since platelets have been reported to modulate immune responses, we aimed to determine the role of platelets in TMEV infection. In transcriptome analyses of platelets, distinct sets of immune-related genes, including major histocompatibility complex (MHC) class I, were up- or downregulated in TMEV-infected mice at different time points. We depleted platelets from TMEV-infected mice by injecting them with platelet-specific antibodies. The platelet-depleted mice had significantly fewer viral antigen-positive cells in the CNS. Platelet depletion reduced the severities of TMEV-IDD and myocarditis, although the pathology scores did not reach statistical significance. Immunologically, the platelet-depleted mice had an increase in interferon (IFN)-γ production with a higher anti-TMEV IgG2a/IgG1 ratio. Thus, platelets may play roles in TMEV infection, such as gene expression, viral clearance, and anti-viral antibody isotype responses. [ABSTRACT FROM AUTHOR]

Ribosomal proteins (r-proteins) are abundant, highly conserved, and multifaceted cellular proteins in all domains of life. Most r-proteins have RNA-binding properties and can form protein–protein contacts. Bacterial r-proteins govern the co-transcriptional rRNA folding during ribosome assembly and participate in the formation of the ribosome functional sites, such as the mRNA-binding site, tRNA-binding sites, the peptidyl transferase center, and the protein exit tunnel. In addition to their primary role in a cell as integral components of the protein synthesis machinery, many r-proteins can function beyond the ribosome (the phenomenon known as moonlighting), acting either as individual regulatory proteins or in complexes with various cellular components. The extraribosomal activities of r-proteins have been studied over the decades. In the past decade, our understanding of r-protein functions has advanced significantly due to intensive studies on ribosomes and gene expression mechanisms not only in model bacteria like Escherichia coli or Bacillus subtilis but also in little-explored bacterial species from various phyla. The aim of this review is to update information on the multiple functions of r-proteins in bacteria. [ABSTRACT FROM AUTHOR]

IntroductionSoil and water salinity has significantly reduced crop yields and threatened human food security worldwide. Understanding the molecular basis of how crop plants receive and respond to salinity and identifying the main components of their stress tolerance are essential for developing tolerant plants through genetic manipulation. Today, functional genomics methods such as transcriptomics, proteomics, and metabolomics have revolutionised the study of plant stress response and facilitated the identification of important pathways and components governing stress tolerance. For example, the production and analysis of EST sequences, RNA microarray, and RNA-sequencing technology (RNA-seq) have made it possible to study and identify gene transcripts in genome-wide stress response networks. Accordingly, this study aimed to analyse the transcript of forage turnip (Brassica rapa L.) under salinity stress, determine the functional orientation of the genome, and identify the important components and genes involved in salinity tolerance.Materials and methodsTwo Expressed Sequenced Tags (ESTs) libraries of forage turnip under non-stress and salinity stress conditions were analysed using bioinformatic and statistical methods. Non-stress and salinity stress libraries had 8408 and 6894 sequences, respectively. After sequences trimming, the genome functional orientation of forage turnip in response to salinity stress was determined based on Fisher's exact test using the DAVID tool. Differentially expressed genes (DEGs) were assigned using the Audio and Claverie statistical test (AC test) implemented in the IDEG6 tool, at a significance level of 5%. Based on Fisher's exact test, a hierarchical gene network among DEGs was predicted. Then, gene network topology analysis was performed using the NetworkAnalyzer plugin in Cytoscape 4.3. One gene was identified as an important gene (Hub gene) based on gene network topology analysis. In a greenhouse experiment using a tolerant genotype and a susceptible genotype of forage turnip, the expression profile of the identified important gene along with some traits related to the plant antioxidant system, including anthocyanin content, antioxidant enzymes activity, and malondialdehyde content, were evaluated in response to different salinity time points. Finally, the relationship among the changes in gene expression, evaluated traits, and salinity tolerance was determined using Correspondence Analysis (CA).Results and discussionPre-processing of 8408 EST sequences of the non-stress library resulted in 8,403 high-quality sequences. Also, out of 6894 EST sequences from the salinity stress library, 6894 high-quality sequences were obtained. BlastX against Arabidopsis proteins showed that 8075 (96.1%) of the non-stress EST sequences had at least one specific hit. 6597 (95.7%) of the EST sequences of the salinity stress library also had at least one specific hit in BlastX. Based on Fisher's exact test, 15 functional groups were significantly (FDR≤0.01) more active in salinity stress conditions than in non-stress conditions. These results clearly showed that under salinity conditions, the genome functional activity of forage turnip was oriented towards response to stresses, especially oxidative stresses, response to internal inductions such as plant hormones, control of metabolic activities, and photosynthetic reactions. Audic and Claverie’s statistical test showed 344 genes were differentially expressed in response to salinity stress; among them, 242 genes were upregulated, and 102 genes were downregulated. The predicted gene network indicated a complex relationship among DEGs, regulatory molecules (especially melatonin and plant hormones), and downstream responsive pathways. Among identified DEGs, the gene encoding transcription factor ZAT6 was assigned as an important gene in the salinity response gene network. The expression profile of the ZAT6 gene, quantity of anthocyanin, the activity of antioxidant enzymes and content of malondialdehyde were significantly different between the two studied genotypes. ZAT6 was significantly more expressed in the stress-tolerant genotype than the stress-susceptible genotype at all time points. In addition, the antioxidant system of the tolerant genotype was more potent than the susceptible genotype. Also, results revealed a significant relationship between the expression profile of ZAT6 and evaluated traits in the context of salinity tolerance. Based on the results, changes in ZAT6 gene expression are directly or indirectly involved in regulating forage turnip plant responses to salinity stress, especially through the control of evaluated traits.ConclusionTranscriptome study clarified some of the molecular bases of the forage turnip response to salinity stress. Accordingly, it seems that the gene encoding the ZAT6 transcription factor plays an important role(s) in the salinity stress tolerance of forage turnip. There was a significant relationship between high expression levels of this gene and enhanced antioxidant activities, which could confirm the hypothesis. However, further studies are needed to assign detailed functions of ZAT6, particularly the association of this gene with the regulatory pathway of melatonin as a major regulatory molecule in plants. This can be an effective starting point for further studies.AcknowledgmentsWe would like to thank the support of the Higher Education Center of Eghlid for this research project.

Climate change is causing alterations in annual temperature regimes worldwide. Important aspects of this include the reduction of winter chilling temperatures as well as the occurrence of unpredicted frosts, both significantly affecting plant growth and yields. Recent studies advanced the knowledge of the mechanisms underlying cold responses and tolerance in the model plant Arabidopsis thaliana. However, how these cold‐responsive pathways will readjust to ongoing seasonal temperature variation caused by global warming remains an open question. In this review, we highlight the plant developmental programmes that depend on cold temperature. We focus on the molecular mechanisms that plants have evolved to adjust their development and stress responses upon exposure to cold. Covering both genetic and epigenetic aspects, we present the latest insights into how alternative splicing, noncoding RNAs and the formation of biomolecular condensates play key roles in the regulation of cold responses. We conclude by commenting on attractive targets to accelerate the breeding of increased cold tolerance, bringing up biotechnological tools that might assist in overcoming current limitations. Our aim is to guide the reflection on the current agricultural challenges imposed by a changing climate and to provide useful information for improving plant resilience to unpredictable cold regimes. Summary statement: In this review, we delve into the molecular mechanisms underlying plant responses to cold. We highlight recent insights into the (epi)genetic pathways controlling cold responses, identifying attractive targets with the potential to address some of the agricultural challenges imposed by climate change. [ABSTRACT FROM AUTHOR]

The transcription factor Oct4 can rightfully be considered a pivotal element in maintaining pluripotency. In addition, its ability to function as a pioneer factor enables the reprogramming of somatic cells back into a pluripotent state. To better understand the regulation of the Oct4-encoding gene (Pou5f1), the main genetic elements that regulate its expression in different states of pluripotency ought to be identified. While some elements have been well characterized for their ability to drive Pou5f1 expression, others have yet to be determined. In this work, we show that translocation of the Pou5f1 gene fragment purported to span all essential cis-elements, including the well-known distal and proximal enhancers (DE and PE), into the Rosa26 locus impairs the self-renewal of mouse embryonic stem cells (ESCs) in the naïve pluripotency state, as well as their further advancement through the formative and primed pluripotency states, inducing overall differentiation failure. These results suggest that regulatory elements located outside the previously determined Pou5f1 boundaries are critical for the proper spatiotemporal regulation of this gene during development, indicating the need for their better characterization. [ABSTRACT FROM AUTHOR]

ACTIN DEPOLYMERIZING FACTOR (ADF) is a conserved protein that regulates the organization and dynamics of actin microfilaments. Eleven ADFs in the Arabidopsis thaliana genome are grouped into four subclasses, and subclass I ADFs, ADF1–4, are all expressed throughout the plant. Previously, we showed that subclass I ADFs function in the regulation of the response against powdery mildew fungus as well as in the regulation of cell size and endoreplication. Here, we report a new role of subclass I ADFs in the regulation of nuclear organization and gene expression. Through microscopic observation of epidermal cells in mature leaves, we found that the size of chromocenters in both adf4 and transgenic lines where expression of subclass I ADF s is downregulated (ADF1-4Ri) was reduced compared with that of wild-type Col-0. Arabidopsis thaliana possesses eight ACTIN (ACT) genes, among which ACT2 , − 7 and − 8 are expressed in vegetative organs. The chromocenter size in act7 , but not in the act2/8 double mutant, was enlarged compared with that in Col-0. Microarray analysis revealed that 1,818 genes were differentially expressed in adf4 and ADF1-4Ri. In particular, expression of 22 nucleotide-binding leucine-rich repeat genes, which are involved in effector-triggered plant immunity, was reduced in adf4 and ADF1-4Ri. qRT-PCR confirmed the altered expressions shown with microarray analysis. Overall, these results suggest that ADF regulates various aspects of plant physiology through its role in regulation of nuclear organization and gene expression. The mechanism how ADF and ACT regulate nuclear organization and gene expression is discussed. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA), the most common form of joint disease, is characterized clinically by joint pain, stiffness, and deformity. OA is now considered a whole joint disease; however, the breakdown of the articular cartilage remains the major hallmark of the disease. Current treatments targeting OA symptoms have a limited impact on impeding or reversing the OA progression. Understanding the molecular and cellular mechanisms underlying OA development is a critical barrier to progress in OA therapy. Recent studies by the current authors' group and others have revealed that the nuclear factor of activated T cell 1 (NFAT1), a member of the NFAT family of transcription factors, regulates the expression of many anabolic and catabolic genes in articular chondrocytes of adult mice. Mice lacking NFAT1 exhibit normal skeletal development but display OA in both appendicular and spinal facet joints as adults. This review mainly focuses on the recent advances in the regulatory role of NFAT1 transcription factor in the activities of articular chondrocytes and its implication in the pathogenesis of OA. [ABSTRACT FROM AUTHOR]

Valproic acid (valproate, VA) has been widely used as an antiepileptic agent for several decades, as well as in the treatment of bipolar affective disorder. For many years, the ability of VA to stop different types of epileptic seizures has been associated with increased GABAergic neurotransmission, inhibition of glutamatergic neurotransmission and a general decrease in CNS hyperactivity by acting on ion channels. However, precise mechanisms of VA anticonvulsant effects are still not clear. Over time, the effects of VA on other neurotransmitter systems, enzymes and intracellular signalling pathways have been revealed, however they do not explain the effectiveness of the drug as an anticonvulsant and mood stabilizer, but only expand its pharmacological profile. Over the past 10 years, the focus of interest in valproate research has changed due to its ability to alter gene expression both by inhibiting histone deacetylases and by changing levels of DNA methylation. These new studies reveal alternative mechanisms of valproate action rather than trying to test and confirm hypotheses previously put forward. [ABSTRACT FROM AUTHOR]

ABSTRACTChlamydia, an obligate intracellular bacterial pathogen, has a unique developmental cycle involving the differentiation of invading elementary bodies (EBs) to noninfectious reticulate bodies (RBs), replication of RBs, and redifferentiation of RBs into progeny EBs. Progression of this cycle is regulated by three sigma factors, which direct the RNA polymerase to their respective target gene promoters. We hypothesized that the Chlamydia-specific transcriptional regulator GrgA, previously shown to activate σ66 and σ28, plays an essential role in chlamydial development and growth. To test this hypothesis, we applied a novel genetic tool known as dependence on plasmid-mediated expression to create Chlamydia trachomatis with conditional GrgA deficiency. We show that GrgA-deficient C. trachomatis RBs have a growth rate that is approximately half of the normal rate and fail to transition into progeny EBs. In addition, GrgA-deficient C. trachomatis fails to maintain its virulence plasmid. Results of RNA-Seq analysis indicate that GrgA promotes RB growth by optimizing tRNA synthesis and expression of nutrient-acquisition genes, while it enables RB-to-EB conversion by facilitating the expression of a histone and outer membrane proteins required for EB morphogenesis. GrgA also regulates numerous other late genes required for host cell exit and subsequent EB invasion into host cells. Importantly, GrgA stimulates the expression of σ54, the third and last sigma factor, and its activator, AtoC, and thereby indirectly upregulating the expression of σ54-dependent genes. In conclusion, our work demonstrates that GrgA is a master transcriptional regulator in Chlamydia and plays multiple essential roles in chlamydial pathogenicity.IMPORTANCEHallmarks of the developmental cycle of the obligate intracellular pathogenic bacterium Chlamydia are the primary differentiation of the infectious elementary body (EB) into the proliferative reticulate body (RB) and the secondary differentiation of RBs back into EBs. The mechanisms regulating these transitions remain unclear. In this report, we developed an effective novel strategy termed dependence on plasmid-mediated expression (DOPE) that allows for the knockdown of essential genes in Chlamydia. We demonstrate that GrgA, a Chlamydia-specific transcription factor, is essential for the secondary differentiation and optimal growth of RBs. We also show that GrgA, a chromosome-encoded regulatory protein, controls the maintenance of the chlamydial virulence plasmid. Transcriptomic analysis further indicates that GrgA functions as a critical regulator of all three sigma factors that recognize different promoter sets at developmental stages. The DOPE strategy outlined here should provide a valuable tool for future studies examining chlamydial growth, development, and pathogenicity.

ABSTRACTThe Plasmodium falciparum alternative histones Pf H2A.Z and Pf H2B.Z are enriched in the same nucleosomes in intergenic euchromatin but depleted from heterochromatin. They occupy most promoters but are only dynamically associated with expression at var genes. In other organisms, acetylation of H2A.Z is important for its functions in gene expression and chromatin structure. Here, we show that acetylated Pf H2A.Z and Pf H2B.Z are dynamically associated with gene expression at promoters. In addition, acetylated Pf H2A.Z and Pf H2B.Z are antagonized by the sirtuin class III histone deacetylases (HDAC) PfSir2A and B at heterochromatin boundaries and encroach upon heterochromatin in parasites lacking PfSir2A or B. However, the majority of acetylated Pf H2A.Z and Pf H2B.Z are deacetylated by class I or II HDACs. Acetylated Pf H2A.Z and Pf H2B.Z are also dynamically associated with promoter activity of both canonical upstream var gene promoters and var gene introns. These findings suggest that both acetylated Pf H2A.Z and Pf H2B.Z play critical roles in gene expression and contribute to maintenance of chromatin structure at the boundaries of subtelomeric, facultative heterochromatin, critical for the variegated expression of genes that enable rapid adaptation to altered host environments.IMPORTANCEThe malaria parasite Plasmodium falciparum relies on variant expression of members of multi-gene families as a strategy for environmental adaptation to promote parasite survival and pathogenesis. These genes are located in transcriptionally silenced DNA regions. A limited number of these genes escape gene silencing, and switching between them confers variant fitness on parasite progeny. Here, we show that PfSir2 histone deacetylases antagonize DNA-interacting acetylated alternative histones at the boundaries between active and silent DNA. This finding implicates acetylated alternative histones in the mechanism regulating P. falciparum variant gene silencing and thus malaria pathogenesis. This work also revealed that acetylation of alternative histones at promoters is dynamically associated with promoter activity across the genome, implicating acetylation of alternative histones in gene regulation genome wide. Understanding mechanisms of gene regulation in P. falciparum may aid in the development of new therapeutic strategies for malaria, which killed 619,000 people in 2021.

ABSTRACTAmong the 16 two-component systems in the opportunistic human pathogen Staphylococcus aureus, only WalKR is essential. Like the orthologous systems in other Bacillota, S. aureus WalKR controls autolysins involved in peptidoglycan remodeling and is therefore intimately involved in cell division. However, despite the importance of WalKR in S. aureus, the basis for its essentiality is not understood and the regulon is poorly defined. Here, we defined a consensus WalR DNA-binding motif and the direct WalKR regulon by using functional genomics, including chromatin immunoprecipitation sequencing, with a panel of isogenic walKR mutants that had a spectrum of altered activities. Consistent with prior findings, the direct regulon includes multiple autolysin genes. However, this work also revealed that WalR directly regulates at least five essential genes involved in lipoteichoic acid synthesis (ltaS): translation (rplK), DNA compaction (hup), initiation of DNA replication (dnaA, hup) and purine nucleotide metabolism (prs). Thus, WalKR in S. aureus serves as a polyfunctional regulator that contributes to fundamental control over critical cell processes by coordinately linking cell wall homeostasis with purine biosynthesis, protein biosynthesis, and DNA replication. Our findings further address the essentiality of this locus and highlight the importance of WalKR as a bona fide target for novel anti-staphylococcal therapeutics.IMPORTANCEThe opportunistic human pathogen Staphylococcus aureus uses an array of protein sensing systems called two-component systems (TCS) to sense environmental signals and adapt its physiology in response by regulating different genes. This sensory network is key to S. aureus versatility and success as a pathogen. Here, we reveal for the first time the full extent of the regulatory network of WalKR, the only staphylococcal TCS that is indispensable for survival under laboratory conditions. We found that WalKR is a master regulator of cell growth, coordinating the expression of genes from multiple, fundamental S. aureus cellular processes, including those involved in maintaining cell wall metabolism, protein biosynthesis, nucleotide metabolism, and the initiation of DNA replication.

Theiler’s murine encephalomyelitis virus (TMEV) infection has been used as a mouse model for two virus-induced organ-specific immune-mediated diseases. TMEV-induced demyelinating disease (TMEV-IDD) in the central nervous system (CNS) is a chronic inflammatory disease with viral persistence and an animal model of multiple sclerosis (MS) in humans. TMEV infection can also cause acute myocarditis with viral replication and immune cell infiltration in the heart, leading to cardiac fibrosis. Since platelets have been reported to modulate immune responses, we aimed to determine the role of platelets in TMEV infection. In transcriptome analyses of platelets, distinct sets of immune-related genes, including major histocompatibility complex (MHC) class I, were up- or downregulated in TMEV-infected mice at different time points. We depleted platelets from TMEV-infected mice by injecting them with platelet-specific antibodies. The platelet-depleted mice had significantly fewer viral antigen-positive cells in the CNS. Platelet depletion reduced the severities of TMEV-IDD and myocarditis, although the pathology scores did not reach statistical significance. Immunologically, the platelet-depleted mice had an increase in interferon (IFN)-γ production with a higher anti-TMEV IgG2a/IgG1 ratio. Thus, platelets may play roles in TMEV infection, such as gene expression, viral clearance, and anti-viral antibody isotype responses.

Ribosomal proteins (r-proteins) are abundant, highly conserved, and multifaceted cellular proteins in all domains of life. Most r-proteins have RNA-binding properties and can form protein–protein contacts. Bacterial r-proteins govern the co-transcriptional rRNA folding during ribosome assembly and participate in the formation of the ribosome functional sites, such as the mRNA-binding site, tRNA-binding sites, the peptidyl transferase center, and the protein exit tunnel. In addition to their primary role in a cell as integral components of the protein synthesis machinery, many r-proteins can function beyond the ribosome (the phenomenon known as moonlighting), acting either as individual regulatory proteins or in complexes with various cellular components. The extraribosomal activities of r-proteins have been studied over the decades. In the past decade, our understanding of r-protein functions has advanced significantly due to intensive studies on ribosomes and gene expression mechanisms not only in model bacteria like Escherichia coli or Bacillus subtilis but also in little-explored bacterial species from various phyla. The aim of this review is to update information on the multiple functions of r-proteins in bacteria.

Recent advances in RNA sequencing technologies helped uncover what was once uncharted territory in the human genome—the complex and versatile world of long noncoding RNAs (lncRNAs). Previously thought of as merely transcriptional "noise", lncRNAs have now emerged as essential regulators of gene expression networks controlling development, homeostasis and disease progression. The regulatory functions of lncRNAs are broad and diverse, and the underlying molecular mechanisms are highly variable, acting at the transcriptional, post-transcriptional, translational, and post-translational levels. In recent years, evidence has accumulated to support the important role of lncRNAs in the development and functioning of the lymphatic vasculature and associated pathological processes such as tumor-induced lymphangiogenesis and cancer metastasis. In this review, we summarize the current knowledge on the role of lncRNAs in regulating the key genes and pathways involved in lymphatic vascular development and disease. Furthermore, we discuss the potential of lncRNAs as novel therapeutic targets and outline possible strategies for the development of lncRNA-based therapeutics to treat diseases of the lymphatic system. [ABSTRACT FROM AUTHOR]

Male sterility is a highly attractive agronomic trait as it effectively prevents selffertilization and facilitates the production of high-quality hybrid seeds in plants. Timely release of mature pollen following anther dehiscence is essential for stamen development in flowering plants. Although several theories have been proposed regarding this, the specific mechanism of anther development in eggplant remains elusive. In this study, we selected an R2R3-MYB transcription factor gene, SmMYB108, that encodes a protein localized primarily in the nucleus by comparing the transcriptomics of different floral bud developmental stages of the eggplant fertile line, F142. Quantitative reverse transcription polymerase chain reaction revealed that SmMYB108 was preferentially expressed in flowers, and its expression increased significantly on the day of flowering. Overexpression of SmMYB108 in tobacco caused anther dehiscence. In addition, we found that SmMYB108 primarily functions as a transcriptional activator via C-terminal activation (amino acid 262–317). Yeast one-hybrid and dual-luciferase reporter assays revealed that genes (SmMYB21, SmARF6, and SmARF8) related to anther development targeted the SmMYB108 promoter. Overall, our results provide insights into the molecular mechanisms involved in the regulation of anther development by SmMYB108. [ABSTRACT FROM AUTHOR]

Herpes simplex virus type-1 (HSV-1) protein ICP27 is an essential immediate early (IE) protein that promotes the expression of viral early (E) and late (L) genes via multiple mechanisms. Our understanding of this complex regulatory protein has been greatly enhanced by the characterization of HSV-1 mutants bearing engineered alterations in the ICP27 gene. However, much of this analysis has been performed in interferon- deficient Vero monkey cells. Here, we assessed the replication of a panel of ICP27 mutants in several other cell types. Our analysis shows that mutants lacking ICP27's amino (N)-terminal nuclear export signal (NES) display a striking cell type-dependent growth phenotype, i.e., they grow semi-permissively in Vero and some other cells but are tightly blocked for replication in primary human fibroblasts and multiple human cell lines. This tight growth defect correlates with a failure of these mutants to replicate viral DNA. We also report that HSV-1 NES mutants are deficient in expressing the IE protein ICP4 at early times postinfection. Analysis of viral RNA levels suggests that this phenotype is due, at least in part, to a defect in the export of ICP4 mRNA to the cytoplasm. In combination, our results (i) show that ICP27's NES is critically important for HSV-1 replication in many human cells, and (ii) suggest that ICP27 plays a heretofore unappreciated role in the expression of ICP4. [ABSTRACT FROM AUTHOR]

Partial root-zone drying (PRD) is an effective water-saving irrigation strategy that improves stress tolerance and facilitates efficient water use in several crops. It has long been considered that abscisic acid (ABA)-dependent drought resistance may be involved during partial root-zone drying. However, the molecular mechanisms underlying PRD-mediated stress tolerance remain unclear. It's hypothesized that other mechanisms might contribute to PRD-mediated drought tolerance. Here, rice seedlings were used as a research model and the complex transcriptomic and metabolic reprogramming processes were revealed during PRD, with several key genes involved in osmotic stress tolerance identified by using a combination of physiological, transcriptome, and metabolome analyses. Our results demonstrated that PRD induces transcriptomic alteration mainly in the roots but not in the leaves and adjusts several amino-acid and phytohormone metabolic pathways to maintain the balance between growth and stress response compared to the polyethylene glycol (PEG)-treated roots. Integrated analysis of the transcriptome and metabolome associated the co-expression modules with PRD-induced metabolic reprogramming. Several genes encoding the key transcription factors (TFs) were identified in these co-expression modules, highlighting several key TFs, including TCP19, WRI1a, ABF1, ABF2, DERF1, and TZF7, involved in nitrogen metabolism, lipid metabolism, ABA signaling, ethylene signaling, and stress regulation. Thus, our work presents the first evidence that molecular mechanisms other than ABA-mediated drought resistance are involved in PRD-mediated stress tolerance. Overall, our results provide new insights into PRD-mediated osmotic stress tolerance, clarify the molecular regulation induced by PRD, and identify genes useful for further improving water-use efficiency and/or stress tolerance in rice. [ABSTRACT FROM AUTHOR]

Genetic transformation of higher eukaryote mitochondria in vivo is an unresolved and important problem. For efficient expression of foreign genetic material in mitochondria, it is necessary to select regulatory elements that provide a high level of transcription and transcript stability. This work is aimed at studying the effectiveness of regulatory elements of mitochondrial genes flanking exogenous DNA using the phenomenon of natural competence of plant mitochondria. For this purpose, genetic constructs carrying the GFP gene under the control of the promoter regions of the RRN26 or COX1 genes and one of the two 3'-untranslated regions (3'-UTR) of mitochondrial genes were imported into isolated Arabidopsis mitochondria, followed by transcription in organello. It was shown that the level of GFP expression under the control of promoters of the RRN26 or COX1 genes in organello correlates with the level of transcription of these genes observed in vivo. At the same time, the presence of the tRNATrp sequence in the 3'-UTR leads to a higher level of the GFP transcript than the presence in this region of the 3'-UTR of the NAD4 gene containing the binding site of the MTSF1 protein. The results we obtained open prospects for creating a system for efficient transformation of the mitochondrial genome. [ABSTRACT FROM AUTHOR]

Selective silencing of 35S rRNA genes (35S rDNA) via nucleolar dominance (ND) is accompanied by epigenetic repatterning, including extensive DNA methylation of promoter sequences through RNA-directed DNA methylation and histone deacetylation. Although ND was first observed nearly a century ago, the molecular mechanisms behind when and how ancestral rDNA sets are chosen for silencing are still unknown. Recent advances in technologies such as single-molecule real-time and nanopore sequencing and optical mapping allow long (>10 kb) DNA segments, including whole rDNA units, to be analysed, which improves characterisation of 35S rDNA loci in plants. Brachypodium hybridum is a unique model species for studying the mechanisms of ND in grasses due to its small nuclear genome size and the presence of only one 35S rDNA locus per subgenome. Nucleolar dominance (ND) is selective epigenetic silencing of 35-48S rDNA loci. In allopolyploids, it is frequently manifested at the cytogenetic level by the inactivation of nucleolar organiser region(s) (NORs) inherited from one or several evolutionary ancestors. Grasses are ecologically and economically one of the most important land plant groups, which have frequently evolved through hybridisation and polyploidisation events. Here we review common and unique features of ND phenomena in this monocot family from cytogenetic, molecular, and genomic perspectives. We highlight recent advances achieved by using an allotetraploid model grass, Brachypodium hybridum, where ND commonly occurs at a population level, and we cover modern genomic approaches that decipher structural features of core arrays of NORs. [ABSTRACT FROM AUTHOR]

Rhizobium tropici CIAT 899 is a broad-host-range rhizobial strain that establishes symbiotic interactions with legumes and tolerates different environmental stresses such as heat, acidity, or salinity. This rhizobial strain produces a wide variety of symbiotically active nodulation factors (NF) induced not only by the presence of plant-released flavonoids but also under osmotic stress conditions through the LysR-type transcriptional regulators NodD1 (flavonoids) and NodD2 (osmotic stress). However, the activation of NodD2 under high-osmotic-stress conditions remains elusive. Here, we have studied the role of a new AraC-type regulator (named as OnfD) in the symbiotic interaction of R. tropici CIAT 899 with Phaseolus vulgaris and Lotus plants. We determined that OnfD is required under salt stress conditions for the transcriptional activation of the nodulation genes and therefore the synthesis and export of NF, which are required for a successful symbiosis with P. vulgaris Moreover, using bacterial two-hybrid analysis, we demonstrated that the OnfD and NodD2 proteins form homodimers and OnfD/NodD2 form heterodimers, which could be involved in the production of NF in the presence of osmotic stress conditions since both regulators are required for NF synthesis in the presence of salt. A structural model of OnfD is presented and discussed.IMPORTANCE The synthesis and export of rhizobial NF are mediated by a conserved group of LysR-type regulators, the NodD proteins. Here, we have demonstrated that a non-LysR-type regulator, an AraC-type protein, is required for the transcriptional activation of symbiotic genes and for the synthesis of symbiotically active NF under salt stress conditions.

microRNA (miRNA) is a type of small non-coding RNA that can participate in cell proliferation and apoptosis by regulating gene expression. More and more evidences show that miRNA-200a is involved in the occurrence and development of non-alcoholic fatty liver disease, alcoholic liver disease, drug-induced liver injury, liver fibrosis, and hepatocellular carcinoma. This article summarizes the differences between miRNA-200a by targeting Downstream target genes of serotonin, regulating related signal pathways and playing different roles in the progression of a variety of liver diseases, provide a reference for exploring the mechanism of a variety of liver diseases. [ABSTRACT FROM AUTHOR]

Transfer RNAs (tRNAs) are well known for their essential function as adapters in delivering amino acids to ribosomes and making the link between mRNA and protein according to the genetic code. Besides this central role in protein synthesis, other functions are attributed to these macromolecules, or their genes, in all living organisms. This review focuses on these extra functions of tRNAs in photosynthetic organisms. For example, tRNAs are implicated in tetrapyrrole biosynthesis, mRNA stabilization or transport, and priming the reverse transcription of viral RNAs, and tRNA-like structures play important roles in RNA viral genomes. Another important function of tRNAs in regulating gene expression is related to their cleavage allowing the production of small non-coding RNAs termed tRNA-derived RNAs. Here, we examine in more detail the biogenesis of tRNA-derived RNAs and their emerging functions in plants. [ABSTRACT FROM AUTHOR]

Abstract Background Heterogeneity of expression quantitative trait locus (eQTL) effects have been shown across gene expression processes. Knowledge on how to produce the heterogeneity is quite limited. This study aims to examine fluctuations in differential gene expression by alleles of sequence variants across expression processes. Results Genome-wide eQTL analyses with transcriptome-wide gene expression data revealed 20 cis-acting eQTLs associated simultaneously with mRNA expression, ribosome occupancy, and protein abundance. A 97 kb-long eQTL signal for mitochondrial ribosomal protein L43 (MRPL43) covered the gene, showing a heterogeneous effect size on gene products across expression stages. One allele of the eQTL was associated with increased mRNA expression and ribosome occupancy but decreased protein abundance. We examined the heterogeneity and found that the eQTL can be attributed to the independent functions of three nucleotide variants, with a strong linkage. NC_000010.11:g.100987606G > T, upstream of MRPL43, may regulate the binding affinity of transcription factors. NC_000010.11:g.100986746C > G, 3 bp from an MRPL43 splice donor site, may alter the splice site. NC_000010.11:g.100978794A > G, in the isoform with a long 3′-UTR, may strengthen the binding affinity of the microRNA. Individuals with the TGG haplotype at these three variants had higher levels of mRNA expression and ribosome occupancy than individuals with the GCA haplotype but lower protein levels, producing the flipped effect throughout the expression process. Conclusions These findings suggest that multiple functional variants in a linkage exert their regulatory functions at different points in the gene expression process, producing a complexity of single eQTLs.

Saline stress is a significant factor that caused crop growth inhibition and yield decline. SHORT INTERNODES/STYLISH (SHI/STY) and SHI-RELATED SEQUENCE (SRS) transcription factors are specific to plants and share a conserved RING-like zinc-finger domain (CX2CX7CX4CX2C2X6C). However, the functions of SHI/STY and SRS genes in cotton responses to salt stress remain unclear. In this study, 26 GhSRSs were identified in Gossypium hirsutum, which further divided into three subgroups. Phylogenetic analysis of 88 SRSs from8 plant species revealed independent evolutionary pattern in some of SRSs derived from monocots. Conserved domain and subcellular location predication of GhSRSs suggested all of them only contained the conserved RING-like zinc-finger domain (DUF702) domain and belonged to nucleus-localized transcription factors except for the GhSRS22. Furthermore, synteny analysis showed structural variation on chromosomes during the process of cotton polyploidization. Subsequently, expression patterns of GhSRS family members in response to salt and drought stress were analyzed in G. hirsutum and identified a salt stressinducible gene GhSRS21. The GhSRS21 was proved to localize in the nuclear and silencing it in G. hirsutum increased the cotton resistance to salt using the virusinduced gene silencing (VIGS) system. Finally, our transcriptomic data revealed that GhSRS21 negatively controlled cotton salt tolerance by regulating the balance between ROS production and scavenging. These results will increase our understanding of the SRS gene family in cotton and provide the candidate resistant gene for cotton breeding. [ABSTRACT FROM AUTHOR]

N‐terminal cysteine oxidases (NCOs) use molecular oxygen to oxidise the amino‐terminal cysteine of specific proteins, thereby initiating the proteolytic N‐degron pathway. To expand the characterisation of the plant family of NCOs (plant cysteine oxidases [PCOs]), we performed a phylogenetic analysis across different taxa in terms of sequence similarity and transcriptional regulation. Based on this survey, we propose a distinction of PCOs into two main groups. A‐type PCOs are conserved across all plant species and are generally unaffected at the messenger RNA level by oxygen availability. Instead, B‐type PCOs appeared in spermatophytes to acquire transcriptional regulation in response to hypoxia. The inactivation of two A‐type PCOs in Arabidopsis thaliana, PCO4 and PCO5, is sufficient to activate the anaerobic response in young seedlings, whereas the additional removal of B‐type PCOs leads to a stronger induction of anaerobic genes and impairs plant growth and development. Our results show that both PCO types are required to regulate the anaerobic response in angiosperms. Therefore, while it is possible to distinguish two clades within the PCO family, we conclude that they all contribute to restrain the anaerobic transcriptional programme in normoxic conditions and together generate a molecular switch to toggle the hypoxic response. Summary statement: Hypoxic induction of plant cysteine oxidases has been acquired and fixed in seed plants from ancestral proteins able to initiate the proteolysis of Cys‐initiating protein substrates by the Arg/N‐degron pathway. [ABSTRACT FROM AUTHOR]

ABSTRACT Histone proteins are found across diverse lineages of Archaea, many of which package DNA and form chromatin. However, previous research has led to the hypothesis that the histone-like proteins of high-salt-adapted archaea, or halophiles, function differently. The sole histone protein encoded by the model halophilic species Halobacterium salinarum, HpyA, is nonessential and expressed at levels too low to enable genome-wide DNA packaging. Instead, HpyA mediates the transcriptional response to salt stress. Here we compare the features of genome-wide binding of HpyA to those of HstA, the sole histone of another model halophile, Haloferax volcanii. hstA, like hpyA, is a nonessential gene. To better understand HpyA and HstA functions, protein-DNA binding data (chromatin immunoprecipitation sequencing [ChIP-seq]) of these halophilic histones are compared to publicly available ChIP-seq data from DNA binding proteins across all domains of life, including transcription factors (TFs), nucleoid-associated proteins (NAPs), and histones. These analyses demonstrate that HpyA and HstA bind the genome infrequently in discrete regions, which is similar to TFs but unlike NAPs, which bind a much larger genomic fraction. However, unlike TFs that typically bind in intergenic regions, HpyA and HstA binding sites are located in both coding and intergenic regions. The genome-wide dinucleotide periodicity known to facilitate histone binding was undetectable in the genomes of both species. Instead, TF-like and histone-like binding sequence preferences were detected for HstA and HpyA, respectively. Taken together, these data suggest that halophilic archaeal histones are unlikely to facilitate genome-wide chromatin formation and that their function defies categorization as a TF, NAP, or histone. IMPORTANCE Most cells in eukaryotic species—from yeast to humans—possess histone proteins that pack and unpack DNA in response to environmental cues. These essential proteins regulate genes necessary for important cellular processes, including development and stress protection. Although the histone fold domain originated in the domain of life Archaea, the function of archaeal histone-like proteins is not well understood relative to those of eukaryotes. We recently discovered that, unlike histones of eukaryotes, histones in hypersaline-adapted archaeal species do not package DNA and can act as transcription factors (TFs) to regulate stress response gene expression. However, the function of histones across species of hypersaline-adapted archaea still remains unclear. Here, we compare hypersaline histone function to a variety of DNA binding proteins across the tree of life, revealing histone-like behavior in some respects and specific transcriptional regulatory function in others.

The transcription factor Oct4 can rightfully be considered a pivotal element in maintaining pluripotency. In addition, its ability to function as a pioneer factor enables the reprogramming of somatic cells back into a pluripotent state. To better understand the regulation of the Oct4-encoding gene (Pou5f1), the main genetic elements that regulate its expression in different states of pluripotency ought to be identified. While some elements have been well characterized for their ability to drive Pou5f1 expression, others have yet to be determined. In this work, we show that translocation of the Pou5f1 gene fragment purported to span all essential cis-elements, including the well-known distal and proximal enhancers (DE and PE), into the Rosa26 locus impairs the self-renewal of mouse embryonic stem cells (ESCs) in the naïve pluripotency state, as well as their further advancement through the formative and primed pluripotency states, inducing overall differentiation failure. These results suggest that regulatory elements located outside the previously determined Pou5f1 boundaries are critical for the proper spatiotemporal regulation of this gene during development, indicating the need for their better characterization.

Phosphorylation of estrogen receptor α (ER) at serine 118 (pS118-ER) is induced by estrogen and is the most abundant posttranslational mark associated with a transcriptionally active receptor. Cistromic analysis of pS118-ER from our group revealed enrichment of the GRHL2 motif near pS118-ER binding sites. In this study, we used cistromic and transcriptomic analyses to interrogate the relationship between GRHL2 and pS118-ER. We found that GRHL2 is bound to chromatin at pS118-ER/GRHL2 co-occupancy sites prior to ligand treatment, and GRHL2 binding is required for maximal pS118-ER recruitment. pS118-ER/GRHL2 co-occupancy sites were enriched at active enhancers marked by H3K27ac and H3K4me1, along with FOXA1 and p300, compared to sites where each factor binds independently. Transcriptomic analysis yielded four subsets of ER/GRHL2-coregulated genes revealing that GRHL2 can both enhance and antagonize E2-mediated ER transcriptional activity. Gene ontology analysis indicated that coregulated genes are involved in cell migration. Accordingly, knockdown of GRHL2, combined with estrogen treatment, resulted in increased cell migration but no change in proliferation. These results support a model in which GRHL2 binds to selected enhancers and facilitates pS118-ER recruitment to chromatin, which then results in differential activation and repression of genes that control estrogen-regulated ER-positive breast cancer cell migration. [ABSTRACT FROM AUTHOR]

CCCTC‐binding factor (CTCF) is an eleven zinc finger (ZF), multivalent transcriptional regulator, that recognizes numerous motifs thanks to the deployment of distinct combinations of its ZFs. The great majority of the ~50,000 genomic locations bound by the CTCF protein in a given cell type is intergenic, and a fraction of these sites overlaps with transcriptional enhancers. Furthermore, a proportion of the regions bound by CTCF intersect genes and promoters. This suggests multiple ways in which CTCF may impact gene expression. At promoters, CTCF can directly affect transcription. At more distal sites, CTCF may orchestrate interactions between regulatory elements and help separate eu‐ and heterochromatic areas in the genome, exerting a chromatin barrier function. In this review, we outline how CTCF contributes to the regulation of the three‐dimensional structure of chromatin and the formation of chromatin domains. We discuss how CTCF binding and architectural functions are regulated. We examine the literature implicating CTCF in controlling gene expression in development and disease both by acting as an insulator and a factor facilitating regulatory elements to efficiently interact with each other in the nuclear space. [ABSTRACT FROM AUTHOR]

CRISPR has revolutionized the way we engineer genomes. Its simplicity and modularity have enabled the development of a great number of tools to edit genomes and to control gene expression. This powerful technology was first adapted to Bacillus subtilis in 2016 and has been intensely upgraded since then. Many tools have been successfully developed to build a CRISPR toolbox for this Gram-positive model and important industrial chassis. The toolbox includes tools, such as double-strand and single-strand cutting CRISPR for point mutation, gene insertion, and gene deletion up to 38 kb. Moreover, catalytic dead Cas proteins have been used for base editing, as well as for the control of gene expression (CRISPRi and CRISPRa). Many of these tools have been used for multiplex CRISPR with the most successful one targeting up to six loci simultaneously for point mutation. However, tools for efficient multiplex CRISPR for other functionalities are still missing in the toolbox. CRISPR engineering has already resulted in efficient protein and metabolite-producing strains, demonstrating its great potential. In this review, we cover all the important additions made to the B. subtilis CRISPR toolbox since 2016, and strain developments fomented by the technology. [ABSTRACT FROM AUTHOR]

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Using a mutant of Mycobacterium smegmatis lacking the major aa3 cytochrome c oxidase of the electron transport chain (Δaa3), we demonstrated that inhibition of the respiratory electron transport chain led to an increase in antibiotic resistance of M. smegmatis to isoniazid, rifampicin, ethambutol, and tetracycline. The alternative sigma factors SigB and SigE were shown to be involved in an increase in rifampicin resistance of M. smegmatis induced under respiration-inhibitory conditions. As in Mycobacterium tuberculosis, SigE and SigB form a hierarchical regulatory pathway in M. smegmatis through SigE-dependent transcription of sigB. Expression of sigB and sigE was demonstrated to increase in the Δaa3 mutant, leading to upregulation of the SigB-dependent genes in the mutant. The pho U2 (MSMEG_1605) gene implicated in a phosphate-signaling pathway and the MSMEG_1097 gene encoding a putative glycosyltransferase were identified to be involved in the SigB-dependent enhancement of rifampicin resistance observed for the Δaa3 mutant of M. smegmatis. The significance of this study is that the direct link between the functionality of the respiratory electron transport chain and antibiotic resistance in mycobacteria was demonstrated for the first time using an electron transport chain mutant rather than inhibitors of electron transport chain. [ABSTRACT FROM AUTHOR]

[Display omitted] • After synthesis mRNAs can be targeted to organelles co-transalationally. • Targeting of mRNAs promotes co-translational folding, assembly and import. • Targeting of mRNAs to organelles enables regulation of gene expression levels. Gene expression is a fundamental and highly regulated process involving a series of tightly coordinated steps, including transcription, post-transcriptional processing, translation, and post-translational modifications. A growing number of studies have revealed an additional layer of complexity in gene expression through the phenomenon of mRNA subcellular localization. mRNAs can be organized into membraneless subcellular structures within both the cytoplasm and the nucleus, but they can also targeted to membranes. In this review, we will summarize in particular our knowledge on localization of mRNAs to organelles, focusing on important regulators and available techniques for studying organellar localization, and significance of this localization in the broader context of gene expression regulation. [ABSTRACT FROM AUTHOR]

The WUSCHEL-related homeobox (WOX) proteins are a class of transcription factors exclusive to plants. They can promote cell division or inhibit stem cell differentiation to regulate plant growth and development. However, the WOX transcription factor genes in the monocotyledon Dendrobium catenatum Lindl. remain relatively uncharacterized. Specifically, the effects of phytohormones on their expression levels are unclear. In this study, we identified and analyzed 10 candidate DcaWOX transcription factor genes in D. catenatum. The DcaWOX family was divided into the modern/WUS, intermediate, and ancient clades. The subcellular localization analysis detected DcaWOX-GFP fusion proteins in the tobacco epidermal leaf cell nucleus. In DcaWOX, members of the WUS clade with the WUS-box motif can significantly activate the expression of TPL in vivo, while members of the intermediate and ancient clades cannot. The expression of the DcaWOX genes varied among the examined tissues. Moreover, the DcaWOX expression patterns were differentially affected by the phytohormone treatments, with differences detected even between homologs of the same gene. Furthermore, the gene expression patterns were consistent with the predicted cis-acting elements in the promoters. The above results suggest that DcaWOX may have an important role in its growth and development and resistance to stress. The results of this comprehensive investigation of the DcaWOX gene family provide the basis for future studies on the roles of WOX genes in D. catenatum. [ABSTRACT FROM AUTHOR]

Cervical cancer (CC) is a preventable disease using proven interventions, specifically prophylactic vaccination, pervasive disease screening, and treatment, but it is still the most frequently diagnosed cancer in women worldwide. Patients with advanced or metastatic CC have a very dismal prognosis and current therapeutic options are very limited. Therefore, understanding the mechanism of metastasis and discovering new therapeutic targets are crucial. New sequencing tools have given a full visualization of the human transcriptome's composition. Non-coding RNAs (NcRNAs) perform various functions in transcriptional, translational, and post-translational processes through their interactions with proteins, RNA, and even DNA. It has been suggested that ncRNAs act as key regulators of a variety of biological processes, with their expression being tightly controlled under physiological settings. In recent years, and notably in the past decade, significant effort has been made to examine the role of ncRNAs in a variety of human diseases, including cancer. Therefore, shedding light on the functions of ncRNA will aid in our better understanding of CC. In this review, we summarize the emerging roles of ncRNAs in progression, metastasis, therapeutics, chemo-resistance, human papillomavirus (HPV) regulation, metabolic reprogramming, diagnosis, and as a prognostic biomarker of CC. We also discussed the role of ncRNA in the tumor microenvironment and tumor immunology, including cancer stem cells (CSCs) in CC. We also address contemporary technologies such as antisense oligonucleotides, CRISPR–Cas9, and exosomes, as well as their potential applications in targeting ncRNAs to manage CC. [ABSTRACT FROM AUTHOR]

Whilst a large number of regulatory mechanisms for gene expression have been characterised to date, transcription regulation in bacteria still remains an open subject. In clinically relevant and opportunistic pathogens, such as Staphylococcus aureus, transcription regulation is of great importance for host-pathogen interactions. In our study we investigated an operon, exclusive to staphylococci, that we name saoABC. We showed that SaoC binds to a conserved sequence motif present upstream of the saoC gene, which likely provides a negative feedback loop. We have also demonstrated that S. aureus ΔsaoB and ΔsaoC mutants display altered growth dynamics in non-optimal media; ΔsaoC exhibits decreased intracellular survival in human dermal fibroblasts, whereas ΔsaoB produces an elevated number of persisters, which is also elicited by inducible production of SaoC in ΔsaoBΔsaoC double mutant. Moreover, we have observed changes in the expression of saoABC operon genes during either depletion of the preferential carbon or the amino acid source as well as during acidification. Comparative RNA-Seq of the wild type and ΔsaoC mutant demonstrated that SaoC influences transcription of genes involved in amino acid transport and metabolism, and notably of those coding for virulence factors. Our results suggest compellingly that saoABC operon codes for a DNA-binding protein SaoC, a novel staphylococcal transcription factor, and its antagonist SaoB. We linked SaoC to the response to nutrient deficiency, a stress that has a great impact on host-pathogen interactions. That impact manifests in SaoC influence on persister formation and survival during internalisation to host cells, as well as on the expression of genes of virulence factors that may potentially result in profound alternations in the pathogenic phenotype. Investigation of such novel regulatory mechanisms is crucial for our understanding of the dynamics of interactions between pathogenic bacteria and host cells, particularly in the case of clinically relevant, opportunistic pathogens such as Staphylococcus aureus. [ABSTRACT FROM AUTHOR]

Background: Heterogeneity of expression quantitative trait locus (eQTL) effects have been shown across gene expression processes. Knowledge on how to produce the heterogeneity is quite limited. This study aims to examine fluctuations in differential gene expression by alleles of sequence variants across expression processes. Results: Genome-wide eQTL analyses with transcriptome-wide gene expression data revealed 20 cis-acting eQTLs associated simultaneously with mRNA expression, ribosome occupancy, and protein abundance. A 97 kb-long eQTL signal for mitochondrial ribosomal protein L43 (MRPL43) covered the gene, showing a heterogeneous effect size on gene products across expression stages. One allele of the eQTL was associated with increased mRNA expression and ribosome occupancy but decreased protein abundance. We examined the heterogeneity and found that the eQTL can be attributed to the independent functions of three nucleotide variants, with a strong linkage. NC_000010.11:g.100987606G > T, upstream of MRPL43, may regulate the binding affinity of transcription factors. NC_000010.11:g.100986746C > G, 3 bp from an MRPL43 splice donor site, may alter the splice site. NC_000010.11:g.100978794A > G, in the isoform with a long 3′-UTR, may strengthen the binding affinity of the microRNA. Individuals with the TGG haplotype at these three variants had higher levels of mRNA expression and ribosome occupancy than individuals with the GCA haplotype but lower protein levels, producing the flipped effect throughout the expression process. Conclusions: These findings suggest that multiple functional variants in a linkage exert their regulatory functions at different points in the gene expression process, producing a complexity of single eQTLs. [ABSTRACT FROM AUTHOR]

Viroids are the smallest pathogens of angiosperms, consisting of non-coding RNAs that cause severe diseases in agronomic crops. Symptoms associated with viroid infection are linked to developmental alterations due to genetic regulation. To understand the global mechanisms of host viroid response, we implemented network approaches to identify master transcription regulators and their differentially expressed targets in tomato infected with mild and severe variants of PSTVd. Our approach integrates root and leaf transcriptomic data, gene regulatory network analysis, and identification of affected biological processes. Our results reveal that specific bHLH, MYB, and ERF transcription factors regulate genes involved in molecular mechanisms underlying critical signaling pathways. Functional enrichment of regulons shows that bHLH-MTRs are linked to metabolism and plant defense, while MYB-MTRs are involved in signaling and hormone-related processes. Strikingly, a member of the bHLH-TF family has a specific potential role as a microprotein involved in the post-translational regulation of hormone signaling events. We found that ERF-MTRs are characteristic of severe symptoms, while ZNF-TF, tf3a-TF, BZIP-TFs, and NAC-TF act as unique MTRs. Altogether, our results lay a foundation for further research on the PSTVd and host genome interaction, providing evidence for identifying potential key genes that influence symptom development in tomato plants. [ABSTRACT FROM AUTHOR]

Filamentous fungi can produce raw-starch-degrading enzyme, however, regulation of production of raw-starch-degrading enzyme remains poorly understood thus far. Here, two novel transcription factors raw-starch-degrading enzyme regulator D (RsrD) and raw-starch-degrading enzyme regulator E (RsrE) were identified to participate in the production of raw-starch-degrading enzyme in Penicillium oxalicum . Individual knockout of rsrD and rsrE in the parental strain Δ ku70 resulted in 31.1%-92.9% reduced activity of raw-starch-degrading enzyme when cultivated in the presence of commercial starch from corn. RsrD and RsrE contained a basic leucine zipper and a Zn2Cys6-type DNA-binding domain, respectively, but with unknown functions. RsrD and RsrE dynamically regulated the expression of genes encoding major amylases over time, including raw-starch-degrading glucoamylase gene PoxGA15A and α-amylase gene amy13A . Interestingly, RsrD and RsrE regulated each other at transcriptional level, through binding to their own promoter regions; nevertheless, both failed to bind to the promoter regions of PoxGA15A and amy13A , as well as the known regulatory genes for regulation of amylase gene expression. RsrD appears to play an epistatic role in the module RsrD-RsrE on regulation of amylase gene expression. This study reveals a novel regulatory pathway of fungal production of raw-starch-degrading enzyme.IMPORTANCETo survive via combating with complex extracellular environment, filamentous fungi can secrete plant polysaccharide-degrading enzymes that can efficiently hydrolyze plant polysaccharide into glucose or other mono- and disaccharides, for their nutrients. Among the plant polysaccharide-degrading enzymes, raw-starch-degrading enzymes directly degrade and convert hetero-polymeric starch into glucose and oligosaccharides below starch gelatinization temperature, which can be applied in industrial biorefinery to save cost. However, the regulatory mechanism of production of raw-starch-degrading enzyme in fungi remains unknown thus far. Here, we showed that two novel transcription factors raw-starch-degrading enzyme regulator D (RsrD) and raw-starch-degrading enzyme regulator E (RsrE) positively regulate the production of raw-starch-degrading enzyme by Penicillium oxalicum . RsrD and RsrE indirectly control the expression of genes encoding enzymes with amylase activity but directly regulate each other at transcriptional level. These findings expand diversity of gene expression regulation in fungi., Competing Interests: The authors declare no conflict of interest.